Polypeptide composition and enzyme activities of the pyrenoid and its regulation by CO2 concentration in unicellular green algae

Abstract

Effects of environmental conditions on the pyrenoid were investigated in unicellular green algae. During adaptation to CO2 limitation, the pyrenoid and pyrenoid starch developed within several hours, while stroma starch was degraded, suggesting that metabolism around the pyrenoid is regulated independently from that in other stromal spaces. This pyrenoid development was light-dependent and inhibited by DCMU but was not affected by changes in nitrogen assimilation. Pyrenoids isolated from Chlamydomonas reinhardtii mainly consisted of the two subunits of ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco), and several other polypeptides were detected. Phosphoribulokinase was not localized in the pyrenoid. Isolated chloroplasts were fractionated to separate intact pyrenoids from stroma and membranes. Rubisco activity was detected in both, and about 70% was distributed in the pyrenoid fraction in low-CO2 cells. The Rubisco content determined by SDS–PAGE, immunoblotting, and immunoelectron microscopy supported the above ratio. By fractionation of isolated chloroplasts under high ionic strength, a pyrenoid-like structure with high Rubisco activity was collected. It is suggested that the main carboxylation site in high-CO2 cells is the stroma, whereas Rubisco is concentrated in the pyrenoid and possibly functions in CO2 fixation under low-CO2 conditions. Key words: Chlamydomonas reinhardtii, chloroplast, CO2 concentration, photosynthetic CO2 fixation, pyrenoid, ribulose 1,5-bisphosphate carboxylase/oxygenase (Rubisco).